Human Immunodeficiency Virus type I (HIV-1) is the etiologic agent of Acquired Immunodeficiency Syndrome (AIDS). AIDS is a complex and multifaceted disease that develops from the infection of cells of the immune system by the HIV virus. The virus has a multi-step life cycle that revolves around the transcriptional control of the virus as regulated by interaction between the viral Tat protein and an RNA element known as the transactivation response (TAR) element. We have recently demonstrated that TAR is utilized by proteins involved in RNA interference (RNAi) and is processed into a viral microRNA (miRNA) that drives remodeling of the viral LTR, leading to heterochromatin formation. The long-term goal of our research is to understand how the RNAi mechanism affects HIV-1 viral replication. RNA interference (RNAi) is a regulatory mechanism conserved in higher eukaryotes. The RNAi pathway generates small interfering RNA (siRNA) or micro RNA (miRNA) from either long double stranded stretches of RNA or RNA hairpins, respectively. The siRNA or miRNA then guides an effector complex to a homologous sequence of mRNA and regulates suppression of gene expression through one of several mechanisms. The HIV-1 TAR element is a stem and loop structure found at the 5'end of all HIV-1 transcripts. The presence of TAR is important for activation of viral transcription by the protein Tat. However, the structure of TAR - a 57 nucleotide hairpin consisting of ~24 basepair stem and a 6 base terminal loop - is very similar to cellular pre- micro RNA (pre-miRNA). These cellular pre-miRNA are acted upon by the enzyme Dicer to yield miRNA that is capable of regulating gene expression. We have recently demonstrated that TAR is cleaved by cellular Dicer to form a viral miRNA. This miRNA is detectable in HIV-1 infected cell lines and primary cells. Further experimental evidence suggests that this viral miRNA may be involved in the induction of transcriptional latency and in the alteration of cellular gene expression. The objective of this project is to determine the mechanism and role of TAR derived miRNA in viral replication. Our hypothesis is that TAR derived miRNA plays a critical role in the viral life cycle as both a mechanism involved in the establishment and maintenance of transcriptional latency and in the alteration of cellular gene expression. Our rationale for this study is based on preliminary data from our lab and the presence of viral miRNA in other human viruses such as human cytomegalovirus, human herpesevirus 8, Epstein Barr virus and peach latent mosaic viroid. The following specific aims will address our hypothesis: (I) What is the mechanism of action of HIV-1 TAR miRNA on viral gene expression? (II) What roles does the viral miRNA play in altering cellular gene expression? Data obtained from these studies will elucidate the role of RNAi in HIV-1 infection and provide new possibilities for inhibiting viral replication. PUBLIC HEALTH RELEVANCE: Currently available drugs for the treatment of HIV-1 do not represent a definitive cure. Research into control of the virus by RNA interference and an increased understanding of the importance of RNAi to the viral life cycle could potentially lead to the discovery of new options in the treatment of HIV/AIDS.